DAPI Research Articles

The impact of graphene oxide nanoparticles on the migratory behavior of metastatic human breast cancer cell, MDA-MB-231.

Breast cancer (BC) with aggressive metastasis is a serious ongoing public health problem among women. Graphene oxide (GO) has an inhibitory effect on the migration rate and metastasis of BC cells, but its various aspects have not yet been explored. This paper aims to research into the effect of GO nanoparticles (GO-Np) on the migratory behavior of MDA-MB-231 as a metastatic human BC cell line. We synthesized GO-Np using a modified Hummer's method. Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), dynamic light scattering (DLS) and zeta potential analyses were conducted to characterize GO-Np. Cytotoxicity of GO-Np against MDA-MB-231 cell lines was examined by optical microscopy and DAPI staining. Migration of cancer cells was analyzed by scratch healing test. Finally, the impact of GO-Np on the expression of BC markers, including Thrombospondin1 (THBS1) and Fibronectin1 (FN1) was investigated by Immunocytochemistry assay (ICC). Results showed a dose-dependent cytotoxicity of GO-Np to MDA-MB-231 cells. The half-maximal inhibitory concentration (IC50) of GO-Np was about 40 μg/ml based on both DAPI test and optical microscopy. The scratch healing test at IC50 concentration of GO-Np, showed a significant decrease (24.4%) in the migration percentage of cells compared to untreated cells. The ICC assay indicated that GO-Np can suppress the expression of both THBS1 and FN1, proving its anti-metastatic properties. In conclusion, GO-Np showed a promising anti-metastatic effect in MDA-MB-231 BC cells.

Lampbrush chromosomes of Danio rerio.

Danio rerio, commonly known as zebrafish, is an established model organism for the developmental and cell biology studies. Although significant progress has been made in the analysis of the D. rerio genome, cytogenetic studies face challenges due to the unclear identification of chromosomes. Here, we present a novel approach to the study of the D. rerio karyotype, focusing on the analysis of lampbrush chromosomes isolated from growing oocytes. Lampbrush chromosomes, existing during diplotene, serve as a powerful tool for high-resolution mapping and transcription analysis due to their profound decondensation and remarkable lateral loops decorated by RNA polymerases and ribonucleoprotein (RNP) matrix. In D. rerio, lampbrush chromosomes are about 20 times longer than corresponding metaphase chromosomes. We found that the lampbrush chromosome stage karyotype of D. rerio is generally undifferentiated, except for several bivalents bearing distinct marker structures, including loops with complex RNP matrix and locus-associated nuclear bodies. Locus-associated nuclear bodies were enriched for coilin and snRNAs; the loci where they formed presumably correspond to the histone gene clusters. Further, we observed the accumulation of splicing factors in giant terminal RNP aggregates on one bivalent. DAPI staining of Danio rerio lampbrush chromosomes revealed large and small chromomeres non-uniformly distributed along the axis. For example, D. rerio lampbrush chromosome 4, comprising the sex-determining region, is divided into two halves-with small chromomeres bearing long lateral loops and with large dense chromomeres bearing no or very tiny lateral loops. As centromeres were not distinguishable, we identified centromeric regions in all bivalents by FISH mapping of pericentromeric RFAL1, RFAL2, and RFAM tandem repeats. Through a combination of morphological analysis, immunostaining of marker structures, and centromere mapping, we developed cytological maps of D. rerio lampbrush chromosomes. Finally, by RNA FISH we revealed transcripts of pericentromeric and telomeric tandemrepeats at the lampbrush chromosome stage.

Enhanced Osteogenic Differentiation for Bone Tissue Engineering: The Synergistic Effect of Adipose‐Derived Mesenchymal Stem Cell Exosomes and Carboxymethyl Chitosan Hydrogel

ABSTRACTExosomes (EXOs) have emerged as critical mediators of intercellular communication with the potential to enhance bone regeneration through paracrine actions. The promising application of EXOs in the tissue engineering (TE) domains has recently sparked heightened research interest. The selection of the type of appropriate biomaterial, along with the method used to produce a scaffold, is another factor determining successful TE. This study assessed the impact of incorporating adipose mesenchymal stem cell‐derived EXOs and its conditioned medium (CM) within a chitosan/carboxymethyl chitosan (CTS/CMC2%) hydrogel on osteogenic differentiation. EXOs were characterized for their morphology, protein content, and size using a scanning electron microscope (SEM), bicinchoninic acid (BCA) protein assay, dynamic light scattering (DLS), and Western blot. Adipose‐derived mesenchymal stem cells (ADMSCs) were encapsulated within hydrogels supplemented with EXOs (12.5 and 25 μg/mL) or CM and tested over 14 days. Cell viability, attachment, and proliferation were enhanced with EXOs and CM treatments versus controls were confirmed by SEM, DAPI staining, and MTT assay. Osteodifferentiation potential was evaluated by alkaline phosphatase activity (ALP), Alizarin‐Red staining (ARS), calcium deposition, and OSTEONECTIN immunostaining (ICC). According to ICC results, encapsulated ADMSCs in the CTS/CMC2% hydrogel treated with 25 μg/mL EXOs (EXO25) exhibited 45.55% OSTEONECTIN expression, the highest percentage observed among all samples. Additionally, encapsulated ADMSCs in CTS/CMC2% hydrogel treated with EXO25 exhibited high ALP expression after 14 days (p < 0.1). In conclusion, EXOs at different concentrations significantly impacted osteogenic differentiation, particularly EXO25, in cooperation with the designed hydrogel (CTS/CMC2%), significantly guiding cell fate for specific targeting differentiation.

2-(4-Bromobenzyl) tethered 4-amino aryl/alkyl-5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidines: design, synthesis, anticancer assessment via dual topoisomerase-I/II inhibition, and in silico studies.

A series of 2-(4-bromobenzyl) tethered 4-amino aryl/alkyl-5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidines (7a-7u) were designed, synthesized, characterized and screened against a panel of cancer cell lines. Compound 7a, in particular, emerged as a potent antiproliferative agent against FaDu cells (HTB-43) with an IC50 value of 1.73 μM. 7a induced morphological alterations in FaDu cells were observed via brightfield microscopy and DAPI staining, confirming cytotoxicity. Autophagy and apoptotic effects of 7a were confirmed by acridine orange staining, Rhodamine 123 staining, and western blot analysis, which revealed dose-dependent increases in LC3A/B and cleaved caspase-3 levels, respectively. Further, 7a impaired cell migration and colony formation, as demonstrated by scratch and clonogenic assays. Additionally, 7a reduced oxidative stress and induced G2/M phase cell cycle arrest in MCF-7 cells. 7a emerged as a dual topoisomerase I and II inhibitor, and results were supported by molecular docking and simulation studies. In anti-inflammatory studies, 7a exhibited selective inhibition of COX-2 over COX-1, supporting its dual anticancer and anti-inflammatory properties.

Bio-fabrication of chitosan-stabilized magnesium oxide nanomaterials: Investigation of photocatalytic, in vitro cytotoxicity activities and apoptosis in oral squamous carcinoma cells.

A bio-fabrication approach is a novel way to develop chitosan-stabilized magnesium oxide nanomaterials (cMgO-NMs). The process involves utilizing polymeric chitosan as the reducing and stabilizing agent. The characteristics of the developed cMgO-NMs were determined using various spectroscopical techniques. Fourier-transform infrared spectroscopy (FTIR) analysis revealed crucial functional groups, Ultraviolet-visible spectroscopy (UV-Vis) spectrum showed nanomaterial development with a peak at 358 nm, and powder X-ray diffraction (PXRD) pattern confirmed a pure cubic crystalline structure. Field emission scanning electron microscopy (FE-SEM) images depicted spherical shape, while energy dispersive X-ray analysis (EDX) confirmed Mg presence. The photocatalytic efficacy of these nanomaterials in degrading dye methylene blue (MB) was examined, and the findings demonstrated the remarkable proficiency of cMgO-NMs in breaking down the dye. The cytotoxic effects of cMgO-NMs were assessed for the first time on PCI-9A and PCI-13 cancer cell lines, yielding an IC50 value of 51 μg/mL and 42 μg/mL. The cMgO-NMs treated PCI-9A and PCI-13 cancer cells morphological changes were observed via acridine orange and ethidium bromide and DAPI staining assay, and apoptotic mode of cell death was examined through flow cytometry and comet assay. Polymeric chitosan proved effective in extensive cMgO-NMs production, showing potential as an anticancer drug, although requiring further preclinical development.

Genome Sequencing Providing Molecular Evidence of Tetrapolar Mating System and Heterothallic Life Cycle for Edible and Medicinal Mushroom Polyporus umbellatus Fr.

Polyporus umbellatus is a species whose sclerotia have been extensively employed in traditional Chinese medicine, which has diuretic, antitumor, anticancer, and immune system enhancement properties. However, prolonged asexual reproduction has resulted in significant homogenization and degeneration of seed sclerotia. In contrast, sexual reproduction has emerged as an effective strategy to address these challenges, with a distinct mating system serving as the foundation for the implementation of sexual breeding. This study presents the first sequencing and assembly of the genome of P. umbellatus, thereby providing an opportunity to investigate the mating system at the genomic level. Based on the annotated mating-type loci within the genome, monokaryotic offspring exhibiting different mating-types were identified. Through the integration of traditional mating tests, the tetrapolar mating system of P. umbellatus was distinctly elucidated. The resequencing of monokaryotic strains with four different mating-types, along with comparative analyses of mating-type loci, revealed the HD1 and HD2 (HD, homeodomain) genes determined the mating A types, and the PR4, PR5, and PR6 (PR, pheromone receptor) genes determined the mating B types. Meanwhile, this study offers a successful case study in the molecular investigation of mating systems. Additionally, the number of sterigma and basidiospores on each basidium was examined using scanning electron microscopy, while the nuclei of basidiospores and basidia at various developmental stages were analyzed through DAPI staining. This research clarifies the heterothallic life cycle of P. umbellatus. The findings of this study are expected to facilitate advancements in genetic research, breeding development, strain improvement, and the industry of P. umbellatus.

Effective Targeting of Glutamine Synthetase with Amino Acid Analogs as a Novel Therapeutic Approach in Breast Cancer

Cancer cells undergo metabolic rewiring to support rapid proliferation and survival in challenging environments. Glutamine is a preferred resource for cancer metabolism, as it provides both carbon and nitrogen for cellular biogenesis. Recent studies suggest the potential anticancer activity of amino acid analogs. Some of these analogs disrupt cellular nucleotide synthesis, thereby inhibiting the formation of DNA and RNA in cancer cells. In the present study, we investigated the anticancer properties of Acivicin and Azaserine in the breast cancer MCF-7 cell line, comparing their effects to those on the non-tumorigenic MCF-10 epithelial cell line in vitro. Interestingly, at lower concentrations, both Acivicin and Azaserine showed potent inhibition of MCF-7 cell proliferation, as assessed by the MTT assay, without detectable toxicity to normal cells. In contrast, Sorafenib (Nexavar), a commonly used drug for solid tumors, showed harmful effects on normal cells, as indicated by increased lactate dehydrogenase (LDH) production in treated cells. Furthermore, unlike Sorafenib, treatment with Acivicin and Azaserine significantly affected apoptotic signaling in treated cells, indicating the role of both amino acid analogs in activating programmed cell death (PCD), as assessed by the Annexin-V assay, DAPI staining, and the relative expression of tumor suppressor genes PTEN and P53. ELISA analysis of MCF-7 cells revealed that both Acivicin and Azaserine treatments promoted the production of anti-inflammatory cytokines, including IL-4 and IL-10, while significantly reducing the production of tumor necrosis factor alpha (TNF-α). Mechanistically, both Acivicin and Azaserine treatment led to a significant reduction in the expression of glutamine synthetase (GS) at both the RNA and protein levels, resulting in a decrease in intracellular glutamine concentrations over time. Additionally, both treatments showed comparable effects on Raf-1 gene expression and protein phosphorylation when compared with Sorafenib, a Raf-1 inhibitor. Moreover, docking studies confirmed the strong binding affinity between Acivicin, Azaserine, and glutamine synthetase, as evidenced by their docking scores and binding interactions with the enzyme crystal. Collectively, these findings provide evidence for the anticancer activity of the two amino acid analogs Acivicin and Azaserine as antagonists of glutamine synthetase, offering novel insights into potential therapeutic strategies for breast cancer.

Semi-quantitative scoring criteria based on multiple staining methods combined with machine learning to evaluate residual nuclei in decellularized matrix

Abstract The detection of residual nuclei in decellularized extracellular matrix (dECM) biomaterials is critical for ensuring their quality and biocompatibility. However, current evaluation methods have limitations in addressing impurity interference and providing intelligent analysis. In this study, we utilized four staining techniques—HE staining, acetocarmine staining, the Feulgen reaction, and DAPI staining—to detect residual nuclei in dECM biomaterials. Each staining method was quantitatively evaluated across multiple parameters, including area, perimeter, and grayscale values, to establish a semi-quantitative scoring system for residual nuclei. These quantitative data were further employed as learning indicators in machine learning models designed to automatically identify residual nuclei. The experimental results demonstrated that no single staining method alone could accurately differentiate between nuclei and impurities. In this study, a semi-quantitative scoring table was developed. With this table, the accuracy of determining whether a single suspicious point is a cell nucleus has reached over 98%. By combining four staining methods, false positives caused by impurity contamination were eliminated. The automatic recognition model trained based on nuclear parameter features reached the optimal index of the model after several iterations of training in 172 epochs. The trained artificial intelligence model achieved a recognition accuracy of over 90% for detecting residual nuclei. The use of multi-dimensional parameters, integrated with machine learning, significantly improved the accuracy of identifying nuclear residues in dECM slices. This approach provides a more reliable and objective method for evaluating dECM biomaterials, while also increasing detection efficiency.

Mycoplasma synoviae Induces Apoptosis in Chicken Oviduct Cells.

Mycoplasma synoviae (MS) is an important pathogen that can cause respiratory diseases in poultry and birds, leading to serious economic losses in the world and impacting the development of the poultry industry. However, the pathogenic mechanisms of MS infection are still unclear, resulting in a lack of effective diagnosis and treatment strategies. This study aimed to uncover the infection effect caused by MS in chicken oviduct cells. MS-infected chicken oviduct cells with different infection times and doses were collected, and DAPI staining was performed to monitor the morphological changes of cell nuclei. Cell viability was detect by CCK-8 assay. The expression of marker genes of cell apoptosis was determined by RT-qPCR and Western blotting assay, respectively. The results showed that MS infection significantly inhibited the growth of chicken oviduct cells, caused the nuclear shrinkage and rupture, activated the expression of apoptosis marker genes Caspase 3 and Beclin-1, and increased the apoptosis rate. These results indicated that MS infection inhibits cell proliferation and promotes apoptosis in chicken oviduct cells, which provided a basis for further revealing the MS pathogenic mechanism and provided a foundation for the future development of anti-infection strategies.

The impact of graphene quantum dots on osteogenesis potential of Wharton's jelly mesenchymal stem cells in fibrin hydrogel scaffolds.

Bone tissue engineering is a promising approach to overcome the limitations of traditional autograft bone transplantation. Graphene quantum dots (GQDs) have been suggested as an enhancement for osteogenic differentiation. This study aimed to investigate the ability of the fibrin hydrogel scaffold in the presence of graphene quantum dots to promote osteogenic differentiation of human Wharton's jelly-derived mesenchymal stem cells (hWJ-MSCs). The hWJ-MSCs were isolated from the Wharton's jelly of the human umbilical cord using a mechanical method. Fibrin hydrogel scaffolds were prepared by mixing 15 µl of thrombin solution with fibrinogen solution. GQDs were incorporated into the scaffolds at concentrations of 0, 5, and 10 µg/ml. Cell viability was determined through DAPI staining and the MTT assay. Osteogenic differentiation was assessed by measuring alkaline phosphatase (ALP) activity, quantifying calcium deposition using Alizarin Red S staining, and analyzing the gene expression of BGLAP, COL1A1, Runx-2 and ALP via qPCR. Scanning electron microscopy (SEM) was employed to analyze the scaffold architecture. SEM analysis revealed that the fibrin hydrogel exhibited a suitable architecture for tissue engineering, and DAPI staining confirmed cell viability. The MTT results indicated that the GQDs and fibrin hydrogel scaffold exhibited no cytotoxic effects. Furthermore, the incorporation of GQDs at a concentration of 10 µg/ml significantly enhanced ALP activity, calcium deposition, and the expression of osteogenesis-related genes compared to the control. The findings suggest that the combination of fibrin hydrogel and GQDs can effectively promote the osteogenic differentiation of hWJ-MSCs, contributing to the advancement of bone tissue engineering.

Phytochemical Profiling and Cytotoxic Potential of Senecio flavus n-Hexane Extract using in vitro and in silico Approaches

Background: Using natural products as potential anticancer and antimicrobial agents has gained significant attention. Methods: This study investigated the anticancer and antimicrobial properties of Senecio flavus by using MTT and well-diffusion assays, respectively. The study also examined the potential of S. flavus extracts to induce apoptosis in liver cancer cells using light and fluorescent microscopy. Gas Chromatography-Mass Spectrometry (GC-MS) was utilized to identify the phyto-compounds, which were subsequently analyzed through molecular docking using AutoDock Vina 1.1.2. Result: The difference in solvent polarity and extraction methods had a notable influence on the yield of the extracts. The saponin fraction had the highest yield (4.42%). Only the hexane extract demonstrated cytotoxic with IC50 values of 241.2 and 187.8 mg/mL for HepG2 and Huh-7 cells, respectively. Apoptotic morphological changes were observed in cells treated with the hexane extract. Additionally, apoptotic cell induction was confirmed using DAPI and AO-EB dual staining. The saponin fraction of S. flavus demonstrated antimicrobial activity, indicated by inhibition zones measuring 19 mm, 27 mm, 13 mm, 17 mm 12 mm against Acinetobacter baumannii (MDR), Acinetobacter baumannii (ATCC® BAA-747), Staphylococcus epidermidis, Staphylococcus aureus Listeria monocytogenes, respectively. GC-MS analysis identified a range of bioactive phyto-compounds within the hexane extract. The compounds (R)-3,5,8a-Trimethyl-7,8,8a,9-tetrahydronaphtho[2,3-b]furan-4(6H)-one and Tricyclo [20.8.0.0 (7,16)]triacontane, 1(22),7(16)-diepoxy- demonstrated the most favourable docking results with 4JAS and 5W1B, respectively, exhibiting a ΔG of -9.7 kcal/mol. This highlights the potential of S. flavus as a promising plant, which requires further investigation and development as a therapeutic agent.

Unveiling Diversity and Function: Venom-Associated Microbes in Two Spiders, Heteropoda venatoria and Chilobrachys guangxiensis

Spiders are natural predators of agricultural pests, primarily due to the potent venom in their venom glands. Spider venom is compositionally complex and holds research value. This study analyzes the diversity of symbiotic bacteria in spider venom glands and venom, as well as the biological activity of culturable symbiotic bacteria. Focusing on the venom glands and venom of Heteropoda venatoria and Chilobrachys guangxiensis, we identified a diverse array of microorganisms. High-throughput sequencing detected 2151 amplicon sequence variants (ASVs), spanning 31 phyla, 75 classes, and 617 genera. A total of 125 strains of cultivable bacteria were isolated. Using the Oxford cup method, crude extracts from 46 of these strains exhibited inhibitory effects against at least one indicator bacterium. MTT (Thiazolyl blue) assays revealed that the crude extracts from 43 strains had inhibitory effects on tumor cell line MGC-803 growth. Additionally, DAPI (4’,6-diamidino-2’-phenylindole) staining and flow cytometry were employed to detect cell apoptosis. The anti-inflammatory activity of nine bacterial strains was assessed using a NO assay kit and enzyme-linked immunosorbent assay (ELISA). This study further investigated the biological activity of venom, exploring the relationship between the venom and the functional activity of venom-associated bacteria.

Initial biological evaluation of nanofiber and microwire biomaterials

Abstract Due to the direct long-term contact with living tissue, the biocompatibility of implants is particularly demanding. In this context, biointegration of implants highly depends on cell colonialization on the surface or in the gap between implant and tissue, as well as cell ingrowth in porous implant structures. Fibrous material is predestined based on two variable parameters, fiber thickness and pore size. Various applications in implant coating, stenting, wound treatment, bone surgery, or cardiovascular field are possible. Coating with cell-friendly material can significantly improve the biocompatibility of the underlying material. In this study, the biocompatibility of medical device samples made of microwire and nanofiber substrates was compared according to ISO 10993-5. In particular, a metal tissue of microwire (stainless steel, 316L) and polymeric nanofiber nonwoven poly-L-lactic acid (PLLA) as well as titanium (Ti) sputtered variants thereof (316L-Ti, PLLA-Ti) were tested in vitro. In order to evaluate their environmental compatibility and their effects on L929 cell line (murine fibroblasts) evaluation was performed via direct contact with 30,000 cells per 48-well in non-adherent plates for 48 h and 96 h. The metabolic activity of cells was determined via CellQuanti-Blue assay, morphological analysis were done using fluorescence staining phalloidin and DAPI. Briefly outlined, microwires were colonized immediately, whereas initially spherical cells settled on nanofiber substrates and later coalesced forming a common cell cluster. The results show independency of structure, ultrathin titanium coating improves cell viability. The findings indicate that the idea of making a surface appealing to cells has potential. It can therefore be assumed that physical vapor deposition (PVD) sputtering of Ti on implant surfaces is a suitable approach to maintain or improve their biocompatibility, regardless of the surface structure of the implants.

Genome content in the non-model ciliate Chilodonella uncinata: insights into nuclear architecture, gene-sized chromosomes among the total DNA in their somatic macronuclei during their development.

Ciliates are a model lineage for studies of genome architecture given their unusual genome structures. All ciliates have both somatic macronuclei (MAC) and germline micronuclei (MIC), both of which develop from a zygotic nucleus following sex (i.e., conjugation). Nuclear developmental stages are not as well explored among non-model ciliate genera, including Chilodonella uncinata (Class- Phyllopharyngea), the focus of our work. Here, we characterize nuclear architecture and genome dynamics in C. uncinata by combining DAPI (4',6-diamidino-2-phenylindole) staining and fluorescence in situ hybridization (FISH) techniques with confocal microscopy. We developed a telomere probe for staining alongside DAPI, which allows for the identification of fragmented somatic chromosomes among the total DNA in the nuclei. We quantify both total DNA and telomere-bound signals to explore changes in DNA content and chromosome maturation across Chilodonella's nuclear life cycle. Specifically, we find that MAC developmental stages in the ciliate C. uncinata are different than the data reported from other ciliate species. These data provide insights into nuclear dynamics during nuclear development and enrich our understanding of genome evolution in non-model ciliates.

Biosynthesis of chitosan encapsulated silver- nanoparticles using Probiotic-Lactobacillus plantarum strain and it's in vitro anticancer assessment on HeLa cells

Cervical cancer remains the deadliest cancer among women worldwide. Investigating the molecular mechanisms of tumor progression by targeting signaling pathways can provide insights into novel therapeutic strategies to overcome the limitations of conventional treatments. Green synthesis of nanoparticles addresses conventional treatment drawbacks like chemotherapy and radiation. This study aims to green synthesize, characterize, and evaluate chitosan-encapsulated silver nanoparticles (AgNPs) using Lactobacillus plantarum probiotics against cervical cancer HeLa cells, targeting the EMT mechanism. The green synthesized chitosan encapsulated Silver- Nanoparticles using probiotic -Lactobacillus plantarum (CS-LP-AgNPs) were characterized using UV–vis spectroscopy, which showed a peak at 420 nm confirming the reduction of AgNPs. Zeta and DLS analysis revealed the particle surface charge and stability. TEM analysis demonstrated that CS-LP-AgNPs are spherically shaped, with a size of approximately 15.3 nm. XRD patterns confirmed the crystalline nature of CS-LP-AgNPs. FTIR spectroscopy confirmed that CS-LP-AgNPs were functionalized with biomolecules. DAPI and double staining were employed to examine characteristic nuclear and morphological changes during apoptosis. Gene expression profiles of EMT in HeLa cell lines were performed to evaluate the anticancer potency of CS-LP-AgNPs. MTT assay demonstrated cytotoxic activity, whereas DAPI/(AO/EB) double staining images showed the induced apoptosis in HeLa cells by CS-LP-AgNPs treatment. CS-LP-AgNPs treated HeLa cells showed decreased SNAIL/EMT via enhancing apoptotic cascade mechanism. Green synthesized CS-LP-AgNPs may be considered an effective anti-cancer drug delivery system for the treatment of CC in the future.

Pretreatment with Notoginsenoside R1 enhances the efficacy of neonatal rat mesenchymal stem cell transplantation in model of myocardial infarction through regulating PI3K/Akt/FoxO1 signaling pathways

BackgroundAlthough stem cell transplantation is a promising approach for the treatment of myocardial infarction (MI), there are still some problems faced such as the low survival rate of stem cells. Here, we investigated the role of Notoginsenoside R1 (NGR1) pretreatment in improving the effects of neonatal rat bone marrow mesenchymal stem cell (MSC) transplantation for treatment of MI.MethodsCardiac functions were detected by echocardiography and the myocardial infarct size was determined by Masson’s trichrome staining in a rat model of MI. The cardioprotective effects of NGR1/LY294002 co-pretreated MSCs was evaluated to explore the underlying mechanism. The angiogenesis was determined by vWF and α-SMA immunofluorescence staining and cell apoptosis was detected by TUNEL. In vitro, the effects of NGR1 on stem cell proliferation was examined by CCK-8 and levels of P-Akt, P-CREB, P-FoxO1 were detected by western blot. Apoptosis, ROS content, and cytokine levels were examined by DAPI and TUNEL staining, a ROS assay kit, and ELISA, respectively.ResultsNGR1 elevated the therapeutic effect of MSC transplantation on infarction by preserving cardiac function, increasing angiogenesis and expressions of IGF-1, VEGF, and SDF-1, and reducing cell apoptosis, whereas the addition of LY294002 prior to NGR1 treatment significantly counteracted the foregoing effects of NGR1. NGR1 pretreatment and SC79 pretreatment were similar in that both significantly increased P-Akt and P-FoxO1 levels in MSC and did not affect P-CREB levels. Besides, both NGR1 and SC79 promoted VEGF, SCF and bFGF levels in MSC cultures, and significantly reduced ROS accumulation and the attenuated cell apoptosis in MSC triggered by H2O2. Similarly, addition of LY294002 before NGR1 treatment significantly counteracted the aforementioned effects of NGR1 in vitro.ConclusionsNGR1 pretreatment enhances the effect of MSC transplantation for treatment of MI through paracrine signaling, and the mechanism underlying this effect may be associated with PI3K/Akt/FoxO1 signaling pathways.

Antiproliferative and Apoptotic Potential of Chicoric Acid, a Major Phytoconstituent of Cichorium intybus, against Colon Cancer Cells: A Possible Inhibition of CDC25 Phosphatases

Background: Over the years, plants have been utilized worldwide for the treatment of cancer in the traditional system of medicine, notably in the majority of developing nations. The use of chemotherapeutic drugs is frequently associated with lethal side effects when treating cancer. Therefore, the panacea to the vicious side effects linked to synthetic pharmaceuticals is the alternative usage of widely accessible and affordable therapeutic approaches. Chicoric acid, a major constituent of Cichorium intybus, has recently been shown to exhibit antioxidant, anti-inflammatory, antidiabetic, antiviral, neuroprotective, hepatoprotective, and anticancer properties. Purpose: Except for a few preliminary studies, no detailed study on the chemotherapeutic efficacy of chicoric acid against cancer cells has been done till now. Therefore, the aim of this study is to assess the potential of chicoric acid against colon cancer cells. Methods: Different in vitro assays like MTT, DAPI staining, rhodamine staining, H2DCFDA staining, caspase activity, and cell analysis were performed on colon cancer HCT116 cells to assess their growth inhibitory properties. Results: According to the results, chicoric acid dose-dependently suppressed the growth of colon cancer HCT116 cells by inducing cell cycle arrest at G0/G1 phase and apoptosis via both intrinsic and extrinsic pathways. In silico findings unveiled the cell cycle regulatory capacity of chicoric acid through binding CDC25 phosphatases, implying its potential as an antiproliferative agent capable of suppressing cell growth. Conclusion: Collectively, the results suggested strong chemopreventive potential of chicoric acid against colon cancer cells. Thus, chicoric acid may turn out to be an excellent anticancer agent, which may be explored in the future for the therapeutic drug development against cancer.

Facile fabrication of dasatinib laden multifunctional polymeric micelles: Evaluation of anti‐proliferative and apoptotic activities in human cancer cells

AbstractDasatinib (DAS) has recently gained significant interest for its anticancer potential. Yet, the lipophilicity inherent in DAS limited its potential use as a chemotherapeutic drug. This study aimed to examine the effectiveness of polyethylene glycol‐polycaprolactone (PEG‐PCL) as a nanocarrier for DAS to increase its anticancer capabilities. The DAS‐loaded PEG‐PCL nanoparticles (termed as DAS@PEG‐PCL NPs) were characterized using Fourier transform infrared (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), and dynamic light scattering (DLS). Morphological staining and MTT tests were employed to investigate drug‐loaded nanoparticles' apoptotic and anti‐proliferative effects. The MTT assay demonstrated that incorporating DAS onto PEG‐PCL NPs resulted in a dose‐dependent increase in cytotoxicity in A549 (lung cancer) and HeLa (cervical cancer) cells. The A549 cancer cells were analyzed for their morphology using the acridine orange/ethidium bromide (AO/EB) and DAPI staining techniques. Overall, these findings demonstrate that the polymeric PEG‐PCL nanoparticle systems hold great potential as a novel therapeutic strategy for cancer treatment.

Shikonin Stimulates Mitochondria-Mediated Apoptosis by Enhancing Intracellular Reactive Oxygen Species Production and DNA Damage in Oral Cancer Cells.

Phytotherapy has rendered a new insight towards the treatment of various cancers, including oral cancer with fewer side effects, over the traditional chemotherapeutic drugs to overcome chemoresistance. Shikonin (Shk) is a natural biologically active alkaloid found in the Lithospermum erythrorhizon plant's root. It has potent cytotoxic activities against various cancers. Our study revealed the release time and anticancer potential of Shk on the SCC9 and H357 oral cancer cell lines. We investigated the antiproliferative, antimigratory, cell cycle arresting and apoptosis promoting activity of Shk in oral cancer cells by performing MTT and morphological assay, colony, and tumor sphere formation assay, AO/EtBr and DAPI staining, Annexin V-FITC/PI staining, assay for reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) measurement, comet assay, qRT-PCR, and western blot analysis. We also checked the interaction of DNA and Shk by docking and CD spectroscopy and EtBr displacement assay. As a result, we found that Shk reduced the viability, proliferation, and tumorigenicity of SCC9 and H357 cells in a time and concentration-dependent manner. We obtained half-maximal inhibitory concentration (IC50) at 0.5 µM for SCC9 and 1.25 µM for H357. It promotes apoptosis via overexpressing proapoptotic Bax and caspase 3 via enhancing ROS that leads to MMP depletion and DNA damage and arrests cells at the G2/M & G2/S phase. The antimigratory activity of Shk was performed by analyzing the expression of markers of epithelial-mesenchymal transition like E-cadherin, ZO-1, N-cadherin, and vimentin. These overall results recommended that Shk shows potent anticancer activity against oral cancer cell lines in both in vitro and ex vivo conditions. So, it could be an excellent agent for the treatment of oral cancer.

A new application of avocado oil to enrich the biological activities of polycaprolactone membranes for tissue engineering.

The metabolites synthesized by plants to protect themselves serves as natural antimicrobial agents used in biomaterials. In this study, avocado oil (AO), was incorporated as a plant source and natural antimicrobial agent into polycaprolactone (PCL) membranes. The effects of varying AO ratios (25, 50, and 100 wt%.-PCL@25AO, PCL@50AO, PCL@100AO) on PCL membrane morphology, chemical structure, wettability, antimicrobial activity, and cell viabilities were investigated. It was demonstrated that the AO acts as a pore-forming agent in solvent-casted membranes. Young's modulus of the membranes varied between 602.68 and 31.92 MPa and more flexible membranes were obtained with increasing AO content. Inhibition zones of AO were recorded between 7.86 and 13.97 mm against clinically relevant microbial strains including bacteria, yeast, and fungi. Antimicrobial activity of AO was retained in PCL membranes at all ratios. Resazurin assay indicated that PCL@25AO membranes were cytocompatible with mouse fibroblast cells (L929 cell line) on day 6 showing 72.4% cell viability with respect to neat PCL membranes. Viability results were supported by scanning electron microscopy images and DAPI staining. The overall results of this study highlight the potential of PCL@25AO membranes as a biomaterial with antimicrobial properties, cytocompatibility, and mechanical strength suitable for various biomedical applications.